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Molecular cloning, expression and polymorphism of goose LRP8 gene. Wang C 2014 et al.
Abstract 1. The low density lipoprotein receptor-related protein 8 (LRP8), a member of the low density lipoprotein receptor (LDLR) gene family, participates in the supplying of lipid during follicular development. The objective of the study was to identify and characterise the LRP8 gene in goose. 2. A 2867 bp fragment that covered the complete coding region (CDS) of goose (Anser cygnoides) LRP8 gene was cloned. It encoded a protein of 917 amino acid residues containing a 24-amino acid signal peptide and 5 functional domains. The goose LRP8 showed high nucleic acid and amino acid identities with those in other species. 3. Similarly to duck LRP8 gene, two splice variants of LRP8, LRP8-1 (containing eight ligand-binding repeats) and LRP8-2 (containing 7 ligand-binding repeats), were identified in goose. 4. Semi-quantitative RT-PCR analysis indicates that the LRP8-1 transcript is expressed in heart, liver, spleen, lung, kidney, breast muscle, duodenum, hypothalamus, pituitary and ovary, negligible or absent in sebum and oviduct, and the LRP8-2 transcript is widely expressed in all examined tissues. 5. A total of 7 SNPs were identified in the coding region of the goose LRP8 gene.
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cloning, expression and association study with reproductive traits of the duck LRP8 gene. Wang C 2013 et al.
Abstract 1. Two splice variants of duck LRP8 were identified, one containing 8 ligand-binding repeats (LRP8-1) and the other containing only 7 repeats (LRP8-2). The two transcripts share ~71-91% nucleic acid identity and ~65-94% amino acid identity with their counterparts in other species. A phylogenetic tree based on amino acid sequences shows that duck LRP8 proteins are closely related to those of chicken, turkey and zebra finch. 2. The semi-quantitative reverse transcription polymerase chain reaction (RT-PCR )analysis indicates that the two transcripts are expressed in all the examined tissues, and the LRP8-1 transcript is more highly expressed in hypothalamus, ovary and pituitary gland than in other detected tissues. 3. Six single nucleotide polymorphisms (SNPs) were identified in the coding region. Association analysis demonstrated that the c.528C > T genotypes were associated with egg production (EP) (EP210d, EP300d and EP360d), age at laying the first egg (AFE) and body weight at sexual maturity (BWSM). The c.1371A > G genotypes were associated with egg production (EP210d, EP300d and EP360d). 4. The haplotypes of c.528C > T and c.1371A?>?G were associated with EP (EP210d, EP300d and EP360d), yolk weight (YW), albumen weight (AW), egg weight (EW), BWSM and the first egg weight (FEW). 5. Duck LRP8 gene was associated with some reproductive traits and is an important candidate gene for the genetic selection of improved reproductive traits.
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Argov N, et al 2004 reported the expression of mRNA of lipoprotein receptor related protein 8, low density lipoprotein receptor, and very low density lipoprotein receptor in bovine ovarian cells during follicular development and corpus luteum formation and regression.
Low-Density Lipoprotein Receptor-Related 8 (LRP8) Is Upregulated in Granulosa Cells of Bovine Dominant Follicle: Molecular Characterization and Spatio-Temporal Expression Studies. Fayad T et al. The low-density lipoprotein (LDL) receptor-related protein 8 (LRP8) is a member of the LDL receptor family that acts in endocytosis and in signal transduction. We cloned the fulllength bovine LRP8 cDNA in granulosa cells (GC) of dominant follicle (DF) as well as several LRP8 mRNA splicing variants, including a variant containing a proline-rich cytoplasmic insert (A(759)-K(817)) involved in intracellular signaling. Expression of the A(759)-K(817) variant was analyzed in GC of follicles at different developmental stages: small follicle (SF; 2-4 mm), DF at day 5 (D5) of the estrous cycle, ovulatory follicles (OF) 24 h after hCG injection, and corpus luteum (CL) at D5. RT-PCR results show that expression is predominant in GC of DF compared to other follicles and CL (P < 0.0001) whereas other related receptors such as LDLR and VLDLR expression does not differ. Temporal analyses of follicular walls from OF following hCG treatment reveal a decrease of LRP8 mRNA expression starting 12 h post-hCG treatment (P < 0.0001). LRP8 protein is exclusively localized in GC with highest levels in DF when compared to SF (P < 0.05). We show that RELN mRNA, encoding an LRP8 ligand, is highly expressed in theca of DF when compared to OF (P < 0.004), whereas MAPK8IP1 mRNA, encoding an LRP8 intracellular interacting partner is expressed in GC of DF. These results demonstrate the differential expression of LRP8, RELN and MAPK8IP1 mRNAs during final follicular growth and ovulation, and suggest that a RELN/LRP8/MAPK8IP1 paracrine interaction may regulate follicular growth.
Lipoproteins in the plasma are the major source of cholesterol obtained by the ovarian theca and granulosa cells for steroidogenesis. In this study, we have identified mRNA expression in bovine theca and granulosa cells of two lipoprotein receptors, low density lipoprotein receptor (LDLr) and very low density lipoprotein receptor (VLDLr) in granulosa cells from small antral follicles through preovulatory follicles and in theca cells from large and medium sized antral follicles. In the corpus luteum (CL) both these receptors were found in the developing and differentiating stages whereas only mRNA for VLDLr was detected in the regression stage. This study also described for the first time, the presence of lipoprotein receptor related protein (LRP8) in granulosa cells from small antral follicles through preovulatory follicles and in theca cells from large and medium sized antral follicles. This may indicate a role of LRP8 in cholesterol delivery to steriodogenic cells. LRP8 was not detected in any of the CL stages. The roles of the LDLr superfamily in lipid transport to ovarian cells and its participation in follicular and CL development and regression is discussed.
Changes in mouse granulosa cell gene expression during early luteinization. McRae RS et al. Changes in gene expression during granulosa cell luteinization have been measured using serial analysis of gene expression (SAGE). Immature normal mice were treated with pregnant mare serum gonadotropin (PMSG) or PMSG followed, 48 h later, by human chorionic gonadotropin (hCG). Granulosa cells were collected from preovulatory follicles after PMSG injection or PMSG/hCG injection and SAGE libraries generated from the isolated mRNA. The combined libraries contained 105,224 tags representing 40,248 unique transcripts. Overall, 715 transcripts showed a significant difference in abundance between the two libraries of which 216 were significantly down-regulated by hCG and 499 were significantly up-regulated. Among transcripts differentially regulated, there were clear and expected changes in genes involved in steroidogenesis as well as clusters of genes involved in modeling of the extracellular matrix, regulation of the cytoskeleton and intra and intercellular signaling. The SAGE libraries described here provide a base for functional investigation of the regulation of granulosa cell luteinization.
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Low-density lipoprotein and cAMP cooperate to regulate expression and function of the low-density lipoprotein receptor-related protein in rat ovarian granulosa cells. Azhar S et al. Rat ovarian granulosa rely heavily on lipoprotein-derived cholesterol for steroidogenesis, which is principally supplied by the LDL receptor- and SR-BI-mediated pathways. In the current study, we characterized the hormonal and cholesterol regulation of another member of the LDL receptor superfamily, LRP and its role in granulosa cell steroidogenesis. Co-incubation of cultured granulosa cells with LDL and Bt2cAMP greatly increased the mRNA/protein levels of LRP. Bt2cAMP and Bt2cAMP + hLDL also enhanced SR-BI mRNA levels. However, there was no change in the expression of RAP, a chaperone for LRP, or another lipoprotein receptor, LRP8/apoER2 in response to Bt2cAMP + hLDL, whereas the mRNA expression of LDL receptor was significantly reduced. The induced LRP was fully functional, mediating increased uptake of its ligand, 2-macroglobulin. The level of binding of another LRP ligand, chylomicron remnants, did not increase, although the extent of remnants degradation that could be attributed to the LRP doubled in cells with the elevated levels of LRP. The addition of lipoprotein type LRP ligands such as chylomicron remnants and VLDL to incubation medium significantly increased the progestin production in both under basal and stimulated conditions. In summary, our studies demonstrate a role for LRP in lipoprotein-supported ovarian granulosa cell steroidogenesis.
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